Carrier polymers migrating into target organs and drug-containing polymers

ABSTRACT

Polymers derived from polymers represented by Formula (A) (d is 20˜500; and Rs, which may be the same or different, represent each H, alkyl or benzyl) by substituting a part or all of the consisting peptide bonds by (1) hydrazino-Glu (Formula B), and saccharide-modified Glu (Formula C) or by (2) hydorazino-Glu (Formula B), and saccharide-modified Glu (Formula C), and drug bonded Glu (Formula D). These polymers, which are carriers optionally bonded to drugs capable of migrating into target organs (cells), are useful as medicines

THE FIELD OF THE ART

The present invention relates to saccharide-modified polymers which areuseful as carriers capable of migrating into target organs (cells),drug-containing polymers using them and the process for the preparationthereof.

BACKGROUND

The drug delivery systems into the target organs comprisinglow-molecular drugs bonded to high-molecular compounds as carrierscapable of migrating into target organs have been studied in order toobtain the aimed pharmaceutical effect of the drugs on the target organsand to reduce the side effects of the drugs on the other organs.

For example, it is disclosed that drug delivery systems into livercomprising of drugs modified with the compounds obtained by combinationof galactose and proteins or high-molecule compounds based on the factthat the receptors specific for galactose exist in liver parenchymalcell in High-Molecule Vol. 46, No. 11, 843-848 (1997).

In addition, it is disclosed that poly-L-glutamic acid derivativeswherein a part of or all of the consisting peptide bonds in thepoly-L-glutamic acid of the formula

(wherein, Xa is degree of polymerization of 20˜540, R^(a) is hydrogen,lower alkyl or benzyl.) are replaced with a group of the formula

is useful as a carrier of drugs capable of migrating into liver in theJapanese Patent Application Kokai Hei 7-228688. The drugs have beenconjugated to carboxyl group in the said glutamic acid via amide bond,ester bond or ion bond etc. directly. Vitamin K5 is described as anexample of drugs in this publication.

Further, it is disclosed that polymers of PGE₁-containing L-glutamicacid derivative (abbreviated as polymer PA) of the formula

is as high-molecule prodrug of PGE₁ capable of migrating into liver inInternational. J. Pharmaceutics, 155, 65-74 (1997). The drug (PGE₁) isconjugated to L-glutamic acid via amide bond through ethylenediamine(—NH—CH₂CH₂—NH—) as a spacer.

In the process for the preparation of the said polymer PA, whichcomprises amidation by condensation between PGE₁ and ethylenediamine asa spacer (reacting the activated ester of PGE₁ with ethylenediamineusing carbodiimide (CDI) etc.), the reaction was carried out in analkaline condition. Therefore, there is a problem that the drug which isunstable in an alkaline condition (e.g. PGE₁) would be decomposed andthat the introducing rate of drugs into poly-L-glutamic acid does notincrease. In this publication, quantity of drugs (PGE₁) introduced intoone molecule of polymer (degree of polymerization of L-glutamicacid=101) is 1.6 molecule.

The present inventors have dissolved such a problem by using hydrazine(—NH—NH—) instead of ethylenediamine (—NH—CH₂CH₂—NH—) as a spacer in thereaction of drugs (e.g. PGE₁) and L-glutamic acid. That is to say, thereaction to introduce the drugs (PGE₁) is carried out in a weak acidiccondition, so it is possible to introduce the drugs constantly, even ifit is unstable in an alkaline condition. Based on this reaction, theyhave improved the introducing rate of drugs (e.g. PGE₁) intopoly-L-glutamic acid, and then succeeded in synthesis ofdrugs-containing polymers showing the superior effect. In addition, ithas proved that any compounds can be introduced into the polymerconstantly by using this reaction. For example, quantity of drugs (PGE₁)introduced into one molecule polymer of the present invention (degree ofpolymerization of L-glutamic acid=97) is 5 molecule, which means thepolymer of the present invention has 3-folds superiority in introducingrate of drug to compare with the polymer of the said publication.

In addition, the polymer using hydrazine of the present invention showssuperiority in both accumulation of drugs into liver afteradministration and effects of drugs (cytoprotective activity of PGE₁) tothe polymers using ethylelendiamine.

Further, there is a merit that such a reaction between hydrazine and thedrug (PGE₁) has been carried out by a simple procedure comprising ofonly stirring them at room temperature.

DISCLOSURE OF THE INVENTION

The present invention relates to

(1) the polymer (abbreviated as Polymer P1.) wherein a part of or all ofthe consisting peptide bonds in the poly-L-glutamic acid of the formula(A)

(wherein, degree of polymerization d is 20˜500, R is hydrogen, C1˜6alkyl or benzyl, with the proviso that each multiple R may be same ordifferent.)

 are replaced with a group of the formula

 wherein

(wherein, G is a modified saccharide capable of conjugating tohydrazine))

as essential substituents with the proviso that when the number ofreplacement groups of the formula (C) is 2 or more, all of said groupsare the same,

(2) the polymer (abbreviated as Polymer P2) wherein a part of or all ofthe peptide bonds in the poly-L-glutamic acid of the formula (A)

 (wherein, all the symbols are defined as hereinbefore)

 (wherein,

 is defined as hereinbefore,

(wherein, D is a drug))

with the proviso that (1) groups of both the formula (C) and (D) areessential substituents, (2) when the number of replacement groups of theformula (C) or (D) is 2 or more, all of said groups of the formula (C)or (D) are the same and (3) the number of replacement groups of theformula (B) may be 0), and (3) the process for the preparation thereof.

DETAILED DESCRIPTION OF THE INVENTION

Polymer P1 is a carrier polymer capable of migrating into target organs(cells) and Polymer P2 is a drug-containing polymer, which is obtainedby utilizing the said carrier polymer, capable of migrating into targetorgans (cells).

The delivery of the polymer of the present invention into target organs(cells) depends upon the saccharide (represented by G) conjugated toglutamic acid. It is known that various kinds of receptors forsaccharides exist in organs (cells) and, new receptors may be found inthe future study. It is possible to obtain the drug delivery system intotarget organs (cells) by choice of saccharide (G) capable of conjugatingto the aimed organs (cells) including such known or new receptors.

For example, in case of monosaccharide, galactose receptor, mannosereceptor and fucose receptor exist in liver parenchymal cells, livernonparenchymal cells (endotherial cells and Kupffer cells) and Kupffercells, respectively, so it is possible to obtain drug delivery systeminto liver (the said liver cells) by conjugate of galactose, mannose orfucose derivative (corresponds to Polymer P1 and P2 of the presentinvention in which

is a group of the formula of (G¹), (G²) and (G³) describedhereinafter.). For example, in case of oligosacchardies such as di, trior tetrasaccharides etc. or multi-saccharides, the delivery of thepolymer of the present invention into target organs (cells) depends uponthe terminal saccharide. For example, the terminal saccharide of lactosewhich is one of disaccharide (corresponds to Polymer P1 and P2 of thepresent invention in which

is a group of the formula (G^(4a)) and (G^(5a)).) is galactose, so sucha polymer migrates into liver parenchymal cell mainly. As for the aimedsaccharide, natural ones or artificial ones which are synthesized may beused.

The symbols and degree of polymerization etc. of Polymer P1 and P2 ofthe present invention are explained in detail as follows:

The symbol d in the formula (A) in Polymer P1 and P2 of the presentinvention means the degree of polymerization of L-glutamic acid which isa unit of the polymer of the present invention and it is an integer of20˜500, preferably 40˜300 and more preferably 50˜150.

The number of replacement of group of the formula (B) in Polymer P1(corresponds to y² described hereinafter.) is 5˜250 and preferably 5˜50.

The number of replacement of group of the formula (C) (corresponds to z²described hereinafter.) is 10˜100, and preferably 20˜60.

The number of replacement of group of the formula (B) in Polymer P2(corresponds to y³ described hereinafter.) is 0˜250, and preferably0˜50.

The number of replacement of group of the formula (C) (corresponds to Z³described hereinafter.) is 10˜100, and preferably 20˜60. The number ofreplacement of group of the formula (D) (corresponds to W³ describedhereinafter.) is 1˜20, and preferably 1˜10.

The average of molecule weight of Polymer P1 is 5,000˜150,000. Forexample, the average of molecule weight of Polymer P1 usingmonosaccharide

is a group of the formula (G¹), (G²) and (G³) described hereinafter.) ordisaccharide such as lactose derivative

is a group of the formula (G^(4a)) and (G^(5a)) described hereinafter.)is 5,000˜100,000 and preferably 10,000˜30,000.

C1-6 alkyl in the formula (A) in Polymer P1 and P2 means methyl, ethyl,propyl, butyl, pentyl or hexyl or its isomer.

Each R is preferably, i) hydrogen or the said C1˜6 alkyl (when multipleR are alkyl, they are the same.), ii) hydrogen or benzyl, or iii)hydrogen only, and more preferably, iii) hydrogen only.

The sacchardie represented by G in the formula (C) in Polymer P1 and P2may be selected in accordance with the receptors which are know or maybe found in the future study exist in the organs (cells), as mentionedbefore. The modified saccharides represented by G capable of conjugatingto hydrazine include, for example, 2-iminoethyl-1-thiosaccharidederivatives and saccharides comprising a group wherein the linkage iscleaved etc.

The said 2-iminoethyl-1-thiosaccharide derivative represented by Ginclude, for example, a group of the formula (if it is represented by

In addition, the said saccharide containing a group, wherein the linkageis cleaved, represented by G include, for example, group of the formula(if it is represented by

(wherein, Q is a saccharide chain containing 1˜10 of saccharide.).

Further, 1˜10 of saccharide represented by Q in the above formulainclude, for example, the saccharide of the formula

(wherein, each p, q and r is 0 or an integer of 1˜9.), preferablygalactose, mannose and fucose (corresponds to a group in which each p,q, r is 0 in the above formula) and more preferably galactose.

is preferably a group of the formula

and more preferably group of the formula (G¹), (G^(4a)) and (G^(5a)).

In Polymer P2, glutamic acid and drugs represented by D is conjugatedvia various kinds of bonds such as hydorazon bond or amide bond etc.through hydrazino (—NH—NH₂) which is introduced to L-glutamic acid inaccordance with the structure of drugs.

The drugs represented by D included any drugs, and preferably, the drugwhich is unstable in an alkaline condition. Such an alkaline conditionmeans pH8˜11 preferably. Of course, it is possible to apply the drugsother than ones which are unstable in an alkaline condition.

Concrete drugs include PGs (e.g. PGEs, PGFs, PGDs), PGIs,naphthyloxyacetic acid derivatives, bicycloalkanoic acid derivatives,guanidinobenzoic acid derivatives, rhodanine acetic acid derivatives,cinnamoic acid derivatives, valproic acid derivatives, Vitamins,anti-allergic agents, anti-vital, anti-cancer agents etc.

PGs include natural PG such as PGE₁, PGE₂, PGF_(1α), PGF_(2α), PGD₁,PGD₂ etc. and its derivatives.

For example, natural PGE₁ and PGE₂ are the compounds shown by thefollowing structures, respectively:

and PGD₁ and PGD₂ are the compounds shown by the following structures,respectively:

The concrete PGs include the compounds of the following formula

(wherein,

is

R^(c) is hydrogen or various kinds of substituents of carboxyl groupsuch as C1˜12 alkyl, benzyl etc.,

A is C2˜10 alkylene (1) in which optional carbon atom may be replacedwith CO and/or (2) may have one or more double bond(s),

B is C1˜10 alkyl, C2˜10 alkenyl or C2˜10 alkynyl may be substituted withphenyl, phenoxy or cycloalkyl (wherein each ring may be substituted withC1˜6 alkyl, C2˜6 alkenyl, C2˜6 alkynyl, C1˜6 alkoxy or halogen etc.),

 is ethylene, trans-vinylene or ethynylene.).

PGs include preferably PGEs or PGDs (the compounds of the formula

in the above formula), more preferably PGEs (the compounds of theformula

in the above formula).

Such compounds include

PGE₁, PGE₂, 17,20-dimethyl-trans-Δ²-PGE₁,6-keto-17,20-dimethyl-trans-Δ²-PGE₁ methyl ester,16,16-dimethyl-trans-Δ²-PGE₁ methyl ester etc.

PGEs and PGDs may be conjugated to L-glutamic acid via hydorazon bond atthe 9th and 11th position carbon, respectively. For example, PGE₁ may beconjugated to L-glutamic acid as shown as following structure:

In addition, PGFs may be conjugated to L-glutamic acid via amide bondbetween the carboxyl group and amine group of hydrazine which isintroduced.

PGIs include natural PGI₂ and its derivatives, for example, thecompounds disclosed in Japanese Patent Application Kokai Sho 54-130543and Sho 55-64541 (corresponding to GBP-2017699). PGls may be conjugatedto L-glutamic acid via amide bond.

Naphthyloxyacetic acid derivatives include, for example, the compoundsdisclosed in Japanese Patent Application Kokai Hei 6-87811(corresponding to U.S. Pat. No. 5,480,998), for example,[5-[2-[1-phenyl-(3-pyridyl)methylildenaminooxy]ethyl]-7,8-dihydronahthalene-1-yloxy]aceticacid shown by the formula

Such a naphthyloxyacetic acid compound may be conjugated to L-glutamicacid via amide bond at the terminal amino group of hydrazine as shown bythe following structure:

Bicycloalkanoic acid derivatives include, for example, the compoundsdisclosed in Japanese Patent Application Hei 9-140959 (corresponding toJapanese Patent Application Kokai Hei 11-29548).

Guanidinobenzoic acid derivatives include, for example, the compoundsdisclosed in Japanese Patent Application Kokai Sho 51-138642(corresponding to U.S. Pat. No. 4,021,472).

Rhodanine acetic acid derivatives include, for example, the compoundsdisclosed in Japanese Patent Application Kokai Sho 57-40478(corresponding to U.S. Pat. No. 4,464,382).

Cinnamoic acid derivatives include, for example, the compounds disclosedin 1) Japanese Patent Application Kokai Sho 55-313 (corresponding toU.S. Pat. No. 4,226,878), 2) Japanese Patent Application Kokai Sho57-131769 (corresponding to U.S. Pat. No. 4,607,046) and 3) WO 98/27053.

Valproic acid derivatives include, for example, the compounds disclosedin Japanese Patent Application Kokai Hei 7-316092 (corresponding toEP-0632008A1).

[The Process for the Preparation of the Polymer of the PresentInvention]

The polymer of the present invention may be prepared by the methoddescribed hereinafter in Examples, known methods or the method of thefollowing reactions (1)˜(3).

(1) introducing of hydrazine to poly-L-glutamic acid,

(2) introducing of saccharide (corresponds to G),

(3) introducing of drugs (corresponds to D).

In the reaction (1), poly-L-glutamic acid of the formula (A)

(wherein, all the symbols are defined as hereinbefore.)

is reacted with hydrazine shown by the formula NH₂—NH₂ in an organicsolvent such as dimethylformadmide (DMF) etc. or without solvent at roomtemperature (10˜25° C.) to prepare the polymer of the formula (II)

(wherein, d¹, x¹ and y¹ are mol (degree of polymerization) of L-glutamicacid connecting COOR¹ (wherein, R¹ is C1˜6 alkyl or benzyl), L-glutamicacid connecting COOH and L-glutamic acid connecting NH₂, respectively.With the proviso that, (1) sum of d¹, x¹ and y¹ equals to d, (2) d¹ maybe 0, (3) each L-glutamic acid connecting COOR¹ (wherein, R¹ is definedas hereinbefore.), COOH and NH₂ may be bonded at random in order.) (seethe method described in J. Appl. Biochem., 2: 25 (1980)).

In the reaction (2), for example, saccharide (G) may be conjugated tohydrazine in the polymer of the formula (II) described hereinafter

(a) by reacting the polymer of the formula (II)

(wherein, all the symbols are defined as hereinbefore.)

 and 2-imino-2-methoxyethyl-1-thiosaccharide in a weak alkalinecondition (e.g. in borate buffer solution (pH9˜10)) or

(b) by reacting the polymer of the formula (II) and various kinds ofsaccharides, and then followed by reduction, if optionally.

2-Imino-2-methoxyethyl-1-thiosaccharide which is the starting materialin the reaction (a) include, for example,2-imino-2-methoxyethyl-1-thiogalactoside,2-imino-2-methoxyethyl-1-thiomanoside or2-imino-2-methoxyethyl-1-thiofucoside of the formula

2-Imino-2-methoxyethyl-1-thiosaccharide is known compound or may beprepared by reacting cyanomethyl-1-thiosaccharide and sodium methoxidein methanol at room temperature (10˜25° C.). (see the method describedin Biochemistry Vol.15, No.18, 3956-3962 (1976)).

The saccharide which is starting material in reaction (b) include, forexample, the compound of the formula

(wherein, Q is defined as hereinbefore.).

The polymer of the present invention wherein the formula

(wherein, Q is defined as hereinbefore.)

may be prepared by reacting aldehyde at the reductive terminal group ofglucose of the saccharide which is used in the reaction and hydrazine inthe polymer of the formula (II). This reaction is carried out in a weakacidic condition (e.g. in citrate buffer solution (pH4˜6)) at roomtemperature (10˜25° C.).

And then, the polymer of the present invention wherein the formula

(wherein, Q is defined as hereinbefore.)

may be prepared by reduction, if optionally.

This reduction is called as reductive amidation. It may be carried outusing reductive agent such as sodium borohydride, sodiumcyanoborohydride etc. in a weak alkaline condition (e.g. in boratebuffer solution (pH8˜9)), at 30˜50° C. By the same procedure, an ordinalsaccharide may be conjugated to hydrazine.

By the known reaction other than the above (a) and (b), saccharide (G)may be conjugated to hydrazine in the polymer of the formula (II).

By the series of the above reactions, the polymer of the presentinvention (corresponds to the said polymer P1) of the formula (I-1)

(wherein, d², x², y² and z² are mol (degree of polymerization) ofL-glutamic acid connecting COOR¹ (wherein, R¹ is defined ashereinbefore.), L-glutamic acid connecting COOH, L-glutamic acidconnecting NH₂ and L-glutamic acid connecting G (saccharide),respectively. With the proviso that (1) sum of d², x², y² and z² equalsto d, (2) d² may be 0, (3) each L-glutamic acid connecting COOR¹(wherein, R¹ is defined as hereinbefore), L-glutamic acid COOH,L-glutamic acid NH₂ and L-glutamic acid G (saccharide) may be connectedat random in order.) may be prepared.

In reaction (3), various kinds of reactions will be carried out inaccordance with the structure of drugs.

1) Drugs possessing keto group (—CO—) may be conjugated via hydorazonbond which is formed by dehydro-condensation reaction with hydrazine inthe polymer of the formula (I-1). This reaction is carried out in a weakacidic condition (e.g. in citrate buffer solution (pH4˜6)), at roomtemperature (10˜25° C.).

2) Drugs possessing carboxyl group (—COOH) may be conjugated via amidebond which is formed by amidation with amino group at the terminal ofhydrazine in the polymer of the formula (I-1). This reaction is wellknown, it may be carried out, for example,

(1) by the method with using acid halide,

(2) by the method with using mixed acid anhydride,

(3) by the method with using conducing agent (EDC and DCC etc.).

3) Besides the above, drugs may be introduced to poly-L-glutamic acidvia various kinds of bonds by known method.

And then, NH₂ in hydrazine in the group may be capped with saccharide byreacting the polymer prepared in reaction (3) and the same saccharide asintroduced in the reaction (2) again, if optionally.

The drug-containing polymer of the present invention (corresponds toPolymer P2) of the formula (I-2)

(wherein, d³, x³, y³, z³ and w³ are mol (degree of polymerization) ofL-glutamic acid connecting COOR¹ (wherein, R¹ is defined ashereinbefore.), L-glutamic acid connecting COOH, L-glutamic acidconnecting NH₂, L-glutamic acid connecting G (saccharide) and L-glutamicacid connecting D(drug). With the proviso that (1) the sum of d³, x³,y³, z³ and w³ equals to d, (2) d³ and y³, independently, may be 0, (3)L-glutamic acid connecting COOR¹ (wherein, R¹ is defined ashereinbefore.), COOH, NH₂, G (saccharide) and D (drug) may be conjugatedat random in order.) may be prepared by series of these reactions.

In each reaction in the present specification, obtained products may bepurified by conventional techniques. For example, purification may becarried out by distillation at atmospheric or reduced pressure, by highperformance liquid chromatography, by thin layer chromatography or bycolumn chromatography using silica gel or magnesium silicate, by washingor by recrystallization. Purification may be carried out after eachreaction, or after a series of reactions.

[Starting Materials and Reagents]

The starting materials and reagents in the present invention are knownper se or may be prepared by known methods.

Industrial Applicability

It has been confirmed that the polymer of the present inventionrepresented as Polymer P1 possesses capability of migrating into targetorgans as shown hereinafter in Experiment. It is expected that the saidpolymer is decomposable in natural condition and that it is safe one,because it is natural high molecule compound. Therefore, the saidpolymer is useful as a carrier.

In addition, it has been confirmed that the drug-containing polymer ofthe present invention represented as Polymer P2 also possessescapability of migrating into target organs and superior effect as shownhereinafter in Experiments.

BEST MODE FOR CARRYING OUT THE INVENTION

The following abbreviations in Experiments and Examples mean as follows:

PLGA: poly-L-glutamic acid,

HZ: hydrazine,

ED: ethylenediamine,

[³H]PGE₁-:

PGE₁ bonded to hydrazine or ethylenediamine wherein the said

PGE₁ is labeled with ³H partially,

Gal: 1-thiogalactpyranosyl-2-imino-ethyl,

-HZ-Lac (reductive): a group of the formula

DMF: dimethylformamide,

MeOH: methanol,

MeONa: sodium methoxide,

EtOH: ethanol.

Experiment 1: Biodistribution of the Carrier Polymer of the PresentInvention (Polymer P1)

PLGA-HZ-Gal (prepared in Example 3) and PLGA-HZ-Lac (prepared in Example5) were labeled with ¹¹¹In and were injected into mouse through its tailvein at dose of 1 mg/kg to analyze biodistribution of them. The resultsare shown in Tables 1 and 2 (Each value in Tables means the percentageof concentration in 1 ml of plasma, the percentage of amount ofaccumulation in each organ and the percentage of urinary excretion ofthe said PLGA derivatives (mean±S.D.), respectively at various timesafter administration.).

TABLE 1 Biodistribution data of PLGA-HZ-Gal 1 min. 5 min. 10 min. 60min. Plasma 47.76 ± 4.87 11.86 ± 4.71 2.41 ± 0.29 0.98 ± 0.16 Kidney3.38 ± 0.41 1.61 ± 0.47 0.75 ± 0.13 0.53 ± 0.17 Spleen 0.02 ± 0.05 0.20± 0.10 0.15 ± 0.04 0.08 ± 0.03 Liver 34.90 ± 0.12 64.80 ± 8.41 75.65 ±3.12 67.40 ± 3.32 Lung 0.11 ± 0.03 0.12 ± 0.01 0.09 ± 0.01 0.06 ± 0.02Urine 0.04 ± 0.06 3.06 ± 4.33 10.93 ± 1.14 15.30 ± 1.11

TABLE 2 Biodistribution data of PLGA-HZ-Lac (reductive) 1 min. 5 min. 10min. 60 min. Plasma 42.16 ± 0.56 17.11 ± 5.85 1.99 ± 2.10 0.06 ± 0.03Kidney 4.49 ± 0.40 4.67 ± 1.15 2.30 ± 1.45 1.24 ± 0.15 Spleen 0.10 ±0.01 0.21 ± 0.02 0.22 ± 0.07 0.19 ± 0.01 Liver 24.53 ± 4.35 44.16 ± 5.4759.51 ± 4.56 56.47 ± 3.57 Lung 0.53 ± 0.10 0.36 ± 0.05 0.11 ± 0.05 0.03± 0.00 Urine 0.16 ± 0.13 5.46 ± 2.89 9.67 ± 3.00 0.91 ± 0.30

About 60% of PLGA-HZ-Gal which was administered was accumulated intoliver at 10 min. after administration. The same level of accumulation ofit was observed in liver at 60 min.

About 60% of PLGA-HZ-Lac which was administered was accumulated intoliver at 10 min. after administration. The same level of accumulation ofit was observed in liver at 60 min.

From the mentioned, it has proved that the carrier polymer of thepresent invention showed high level of accumulation and long-termaccumulation of it in liver.

Experiment 2: Biodistribution of the Drug-containing Polymer of thePresent Invention (Polymer P2)

Biodistribution of [³H]PGE₁-HZ-PLGA-HZ-Gal (prepared in Example 4,degree of polymerization=97) and [³H]PGE₁-ED-PLGA-ED-Gal (Comparison:the polymer described in International J. Pharmaceutics, 155, 65-74(1997), degree of polymerization=101) was analyzed by the same procedureas described in Experiment 1. The results are shown in Tables 3(Invention) and 4 (Comparison) (Each value in Tables means thepercentage of concentration in 1 ml of plasma, the percentage of amountof accumulation in each organ and the percentage of urinary excretion ofthe said [³H]PGE₁ derivatives (mean±S.D.), respectively at various timesafter administration.).

TABLE 3 Biodistribution data of [³H]-PGE₁-HZ-PLGA-HZ-Gal 1 min. 5 min.10 min. 60 min. Plasma 14.33 ± 0.75 1.64 ± 0.51 0.40 ± 0.06 0.26 ± 0.07Kidney 1.26 ± 0.23 1.23 ± 0.06 0.79 ± 0.14 0.64 ± 0.22 Spleen 1.11 ±0.14 1.85 ± 0.10 1.24 ± 0.17 1.84 ± 0.16 Liver 54.42 ± 0.79 70.39 ± 3.5180.54 ± 9.52 85.43 ± 3.78 Lung 1.88 ± 0.46 1.64 ± 0.56 1.00 ± 0.21 0.44± 0.18 Urine 0.00 ± 0.00 1.17 ± 1.01 1.79 ± 0.25 1.45 ± 0.83

TABLE 4 Biodistribution data of [³H]-PGE₁-ED-PLGA-ED-Gal 1 min. 5 min.10 min. 60 min. Plasma 36.55 ± 1.63 6.33 ± 0.84 1.99 ± 0.42 0.00 ± 0.00Kidney 9.58 ± 1.46 28.82 ± 2.82 33.25 ± 5.63 13.16 ± 1.32 Spleen 0.41 ±0.06 0.37 ± 0.17 0.63 ± 0.27 1.02 ± 0.53 Liver 27.72 ± 3.81 41.16 ± 2.0447.19 ± 1.03 45.12 ± 8.21 Lung 1.97 ± 0.85 1.40 ± 0.13 0.82 ± 0.33 0.69± 0.14 Urine 0.05 ± 0.05 3.77 ± 2.28 2.28 ± 1.61 6.56 ± 3.15

As shown in Table 3, 70% of drug which was administered was accumulatedinto liver at 5 min. after administration. In addition, 85% and 70% ofdrug were observed to be accumulated to liver at 1 hour and 24 hoursafter administration, respectively.

On the other hand, in Comparison (Table 4) group, 40% and 45% of drugwhich was administered were accumulated to liver at 5 min. and 1 hourafter administration, respectively.

Therefore, it has proved that it is possible to deliver the drug at thehigher concentration continuously into liver using the drug-containingpolymer of the present invention.

Experiment 3: Effect of the Drug (PGE₁)-containing Polymer of thePresent Invention (Polymer P2) on CCl₄ Induced Liver Damage

A solution of 10% (v/v) of CCl₄ in sesame oil at dose of 10 ml/kg wasadministered into mouse abdominal cavity, and then drug (saline solution(Control), Free-PGE₁ (Comparison), drug (PGE₁)-containing polymer of thepresent invention PGE₁-HZ-PLGA-HZ-Lac (reductive) (prepared in Example6)) were injected into mouse through its tail vein at the setting dose.After the mouse had been fasted for 18 hours (25° C., water was freelygiven), blood was collected to assay GPT level (IU/L) in plasma. Theresults are shown in Table 5.

TABLE 5 n (No. of animals) GPT level Control (saline solution/CCl4 (−))3 12.68 ± 1.527 Control (saline solution/CCl4 (+)) 5 614.56 ± 250.3 Free PGE₁ (0.065 mg/kg) 5 660.89 ± 218.28 PGE₁-HZ-PLGA-HZ-Lac 4 239.12 ±77.482 (1 mg/kg)

As shown in Table 5, the drug (PGE₁)-containing polymer of the presentinvention showed inhibition effect on increasing GPT level in plasma ofCCl₄ induced liver damage significantly to compare with the groupconsisting of saline solution (Control group). In addition, theinhibition rate of increasing GPT level in the Invention group wasthree-time superior to that in the group consisting of free PGE₁ at thecorresponding dose.

REFERENCE EXAMPLE AND EXAMPLES

The following Reference Examples and Examples are intended toillustrate, but not limit, the present invention. Each numberrepresented as d^(t), x^(t), y^(t) (t=1, 2, 3), z^(u) (u=2, 3), w³ inthe column of degree of polymerization means mol of L-glutamic acidconnecting COOR¹ (wherein, R¹ is C1˜6 alkyl, benzyl.), L-glutamic acidconnecting COOH, L-glutamic acid connecting NH₂, L-glutamic acidconnecting G (galactose form or lactose form) and L-glutamic acidconnecting D (PGE₁) per 1 mol of polymer.

Reference Example 1

Synthesis of PLGA-HZ (Degree of Polymerization: d¹=0, x¹=29, y¹=50)

To γ-benzyl-poly-L-glutamic acid (MW: 17,300, degree ofpolymerization=79) (200 mg), solution of hydrazine.monohydrate (10 ml)in DMF (3 ml) was added at a dropwise with stirring. The mixture wasreacted for 3 hours at room temperature. The reaction solution wasdialyzed with dialysis tube (3,500 molecular weight cut-off) (When theinner solution of tube became to be gel, the solution was recovered tobe homogenous condition by addition of an adequate quantity of conc.HCl.). Inner solution of tube was ultrafiltered (10,000 molecular weightcut-off), concentrated and freezed to dry to obtain the title compoundhaving the following physical data.

It was confirmed that each benzyl group, which was a protecting group ofglutamic acid, was removed entirely by NMR analysis. In addition,hydrazine residue was assayed by β-naphathoquinon-4-sulphonate method.

MW: 10,900; degree of polymerization: d¹=0, x¹=29, y¹=50.

Example 1

Synthesis of PLGA-HZ-Gal (Degree of Polymerization: d²=0, x²=29, y²=8,z²=42)

(1) To cyanomethyl 1-thiogalacoside (150 mg), MeONa/MeOH (3 ml) wasadded. The mixture was stirred for 24 hours. MeOH was distilled offunder reduced pressure from the mixture.

(2) PLGA-HZ (prepared in Reference Example 1) (50 mg) was dissolved in2N HCl (1 ml). The mixture was neutralized by addition of 2N NaOH.Borate buffer solution (50 mM, pH9.5)(3 ml) was added thereto. Thesolution was added to the residue obtained in (1). The mixture wasstirred for 5 hours at room temperature. The reaction solution wasdialyzed, concentrated and freezed to dry to obtain the title compoundhaving the following physical data. In addition, Gal residue was assayedby sulphate-anthron method.

MW: 20,900; degree of polymerization: d²=0, x²=29, y²=8, z²=42.

Example 2

Synthesis of [³H]PGE₁-HZ-PLGA-HZ-Gal (Degree of Polymerization: d³=0,x³=29, y³=7, z³ =42, w ³=1)

(1) PLGA-HZ-Gal (prepared in Example 1) (22.5 mg) was dissolved in 0.1Macetate buffer solution (pH5.0) (1 ml).

(2) To a solution of iced PGE₁ (2.5 mg) in EtOH (0° C., 1 ml), asolution of [³H]PGE₁ (EtOH: H₂O=7:3; 0.5 μCi/ml) (0.1 ml) was added.

(3) Stirring the solution prepared in the above (1) at room temperature,the solution obtained in the above (2) was added at a dropwise thereto.0.1M acetate buffer solution (pH5.0) (0.5 ml) was added thereto toclarify the solution. The solution was stirred for 24 hours at 4° C.After removing the impurities from the reaction mixture, the solutionwas dialyzed. The dialyzed solution was ultrafiltered (10,000 molecularweight cut-off, concentrated and freezed to dry to obtain the titlecompound having the following physical data.

MW: 21,200; degree of polymerization: d³=0, x³=29, y³=7, z³=42, w³=1.

Example 3

Synthesis of PLGA-HZ-Gal (Degree of Polymerization: d²=0, x²=29, y²=37,z²=31)

By the same procedure as Reference Example 1→Example 1, the titlecompound having the following physical data was obtained usingγ-benzyl-poly-L-glutamic acid (degree of polymerization=97).

MW: 20,800; degree of polymerization: d²=0, x²=29, y²=37, z²=31.

Example 4

Synthesis of PGE₁-HZ-PLGA-HZ-Gal and [³H]PGE₁-HZ-PLGA-HZ-Gal (Degree ofPolymerization: d³=0, x³=29, y³=32, z³=31, w³=5)

PLGA-HZ-Gal (prepared in Example 3) (20 mg) was dissolved in 0.01Macetate buffer solution (pH5.0) (5 ml). Stirring this solution, asolution of PGE₁ (4 mg) in EtOH (0.5 ml) was added at a dropwisethereto. The mixture was stirred over night at room temperature. Thereaction solution was dialyzed by saline solution to obtain the titlecompound (PGE₁-HZ-PLGA-HZ-Gal) having the following physical data. Inaddition, by the same procedure, the title compound ([³H]PGE₁) havingthe following same physical data was obtained by addition of [³H]PGE₁(10μCi) to the said solution of PGE₁ in EtOH. Both compounds were stored asa solution form.

MW: 23,000; degree of polymerization: d³=0, x³=29, y³=32, z³=31, w³=5.

Example 5

Synthesis of PLGA-HZ-Lac (Reductive/degree of Polymerization: d²=0,x²=35, y²=40, z²=22)

PLGA-HZ (MW: 13,300, degree of polymerization: d¹=0, x¹=35, y¹=62) (50mg) which was prepared by the same procedure as Reference Example 1using γ-benzyl-poly-L-glutamic acid (degree of polymerization=97) wasdissolved in 5N NaOH and neutralized to about pH7 by addition of 5N HCl.0.1M borate buffer solution (pH8.5) was added thereto to become topH8˜9. Lactose (143 mg) and sodium cyanoborohydride (50 mg) was addedthereto. The solution was reacted for one day at 37° C. The reactionsolution was purified with dialysis and freezed to dry to obtain thetitle compound having the following physical data.

MW: 20,800; degree of polymerization: d²=0, x²=35, y²=40, z²=22.

Example 6

Synthesis of PGE₁-HZ-PLGA-HZ-Lac (Reductive/degree of Polymerization:d³=0, x³=35, y³=36, z³=22, w³=4)

By the same procedure as Example 2, the title compound having thefollowing physical data was obtained using PLGA-HZ-Lac(reductive/prepared in Example 5).

MW: 22,800; degree of polymerization: d³=0, x³=35, y³=36, z³=22, w³=4.

What is claimed is:
 1. A polymer of the formula (A-1):

wherein d is an integer of 20-500, each X is independently selected fromthe group consisting of —OR, —NH—NH₂ and

wherein each R is independently selected from the group consisting ofhydrogen, C1-6 alkyl and benzyl, and wherein

wherein G is a saccharide, with the proviso that (1) at least one X is—NH—NH₂ and at least one X is

 and (2) when more than one X is

 all of the

 groups are the same.
 2. A polymer according to claim 1, wherein

is

wherein, Q is a saccharide chain containing 1-10 saccharide subunits. 3.The polymer according to claim 1, wherein

is